Anchorage dependent and non-anchorage dependent growth of cells notes

 Anchorage dependent and non-anchorage dependent growth of cells

Anchorage dependent growth of cells:

1. Cells which require attachment(support) for their growth and multiplication are are said to be anchorage dependent growers.
2. Majority of vertebrate cells cultured in vitro have been grown as monolayers on an artificial supporting substrates like glass, plastic, metalic surfaces, semi-solid substrates, feeder layer etc.
3. most cells need to spread out on a substrate in order to proliferate, inadequate spreading will inhibit cell multiplication.
4. Most of the mono layer primary cultures or continuous cell lines need a periodic change of medium whether or not the cell are proliferating. In cultures where cells are proliferating, the usual practice in sub culturing an adherent cell line involves following steps: i) Cultivation of animal cells start by dissociation of tissues into fragments or isolated cells obtained by treating with proteolytic enzyme or mechanical method. Then the dispersed cells are placed in a plastic container containing liquid medium. After a log period the cells attach themselves to the bottom of the container and start dividing mitotically to produce mono layer as primary cell lines. ii) For sub-culturing, remove medium and add PBSA along the sides of the flask opposite the cells to avoid dislodging of cells, rinse the cells and discard the PBSA. This is designed to remove traces of serum that would inhibit the action of trypsin. iii) Dissociate the cells round up(5-15min), when the bottle is tilted and the mono layer slide down the surface. iv) Add the medium(0.1-0.2ml/cm2) and dispense cells by repeated pipetting over the surface bearing the mono-layer to dispense the cell line, pipette the cell suspension up and down for few minutes to get single cell suspension. v) A single cell suspension is desirable at subculture to ensure an accurate cell count and uniform growth. It is necessary where cells are to be isolated as clones.
5. Intervals between change of medium and sub-culturing vary from one cell line to another depending upon the rate of growth of metabolism. Rapidly growing cell lines like "Hela" are sub-cultured once per week and medium change every four days more slowly growing cell lines are sub-cultured every 2 or four weeks and the medium changed every week between subcultures. There are several factors which are used to assess the need for replacement of medium these include i) drop in pH, ii) Cell conc. iii) cell type and iv) cell morphology etc.
6. Sources of adherent cells derived from such organs in body that there also they are not motile example; nerve cells, liver, kidney, muscle etc.

Non-anchorage dependent growth of cells:

1. Generally normal cell lines propogate in unaltered form for a limited number of cell generations, beyond which they may either die or give rise to continous cell lines because certain primary cell lines undergo spontaneous transformation and frequently become non-anchorage dependent hence grow in suspension.
2. Alteration in the phenotypic property in a normal cell culture may also be due to  chemical or viral induction.
3. continuous cell lines often aneuploid and prefer to grow in suspension. They found to over grow and become immortal cells. The density of division may increase in such a eay that the cells grow in a clumps rather than in monolayer. These cells may be irregularly oriented with respect to each other.
4. No enzyme treatment is required in this case and the sub-culturing is done by dilution of cell density, rather than by replacement of the medium because it will require centrifugation of the suspension culture to obtain a pellet of cells to be added to a fresh quantity of the medium.